Nucleic acid bioassays that can be performed on-site or at point-of-care (POC) with minimal equipment, rapidly and at low cost are in high demand. While there are many attempts aimed at achieving this, none are in practice currently. Techniques and methods for detecting disease DNA biomarkers are known, including the polymerase chain reaction (PCR) and ligase chain reaction (LCR)1. However, these methods require thermocycling on a thermal cycler to achieve rapid exponential DNA amplifications and hence are not suitable for field or on-site applications. Nonetheless, various isothermal DNA amplification methods have emerged recently to overcome this limitation2,3. For instance, to enable POC applications, pathogen DNA amplified by the recombinase polymerase amplification (RPA)4 or helicase dependent amplification (HDA)5 have been adapted for detection on lateral flow strips and portable fluorometers6-9. However, such readout methods whilst convenient, are still dependent on the use of relatively sophisticated equipment and may still present a financial obstacle for operators worldwide. Moreover, on-field sampling is rarely, if ever, discussed. In particular, consistently generating a fixed amount of sample DNA with minimal human manipulation and on-site infrastructure for downstream applications. This non-trivial issue may have significant impact on the performance of any assay. Therefore, a field-ready comprehensive assay for on-site nucleic acid detection applications is still an elusive aspiration.
One area that can benefit from low cost on-site assays is in agriculture. The agricultural industry is a major contributor to the world economy (estimated as $1500 billion per year). However, crop disease outbreaks are major issues in agriculturally reliant economies, especially in developing countries, with estimated yearly crop losses of $220 billion10. Currently, there is no way to salvage compromised crops once a disease outbreak spreads beyond a certain threshold. The ideal method to control disease outbreaks is by early detection in the field before it spreads. The effectiveness of crop disease management is highly dependent on the rapidness, sensitivity and specificity of the diagnostic method. Disease identification is traditionally done by visual examination of the disease phenotype in affected plant tissues. However this requires experienced plant pathologists and is relatively subjective11. Many sensitive diagnostic methods to enhance disease identification such as enzyme-linked immunosorbent assays (ELISAs)12,13, immunoblots14,15, immunofluorescent tests16 and various iterations of PCR based assays17,18 have been developed to facilitate disease diagnosis. However, all these detection methods require expensive and sophisticated equipment and can only be performed in laboratories by well-trained technicians. Moreover, the lack of rapid on-site detection can lead to delays in the deployment of disease control measures, which in turn, leads to further crop losses. It is therefore essential to develop new disease diagnostic technologies that can be applied in the field without the need to access specialized laboratory equipment. In addition, these technologies should be cheap, sensitive, reproducible and require no specialized personnel with the ultimate goal of allowing each farmer to monitor his or her own crops. Similarly, early detection of farm animal pathogens is essential to avoid the spread of diseases, especially in modern farms using intensive production methods. In addition, early diagnosis and detection of human diseases and the availability of POC methods nondependent on sophisticated technological requirements can save innumerable lives in developed as well as developing countries particularly after natural disaster situations such as typhoons, tsunamis or earthquakes.